Drug resistant Gram-negative bacterial pathogens are an increasing cause of hospital-acquired infections, mortality, and a huge burden on healthcare costs. Acinetobacter baumannii is a major cause of such infections and strains have recently emerged which are resistant even to the last line of defense drugs, polymyxin B and colistin (polymyxin E), which target and disrupt the lipid A portion of lipopolysaccharide (LPS) in the outer membrane. Understanding the mechanism of this resistance at the molecular level would facilitate the development of novel therapeutics aimed at reversing resistance, in much the same way that beta-lactamase inhibitors can counteract penicillin resistance. To this end, we have recently identified a novel protein that we have named ArmR (or Armor - Antimicrobial Resistance by Modification of lipid A surface chaRge) that is widely conserved and which we show is required for resistance to polymyxins in A. baumannii as well as other Gram-negative bacteria. ArmR is required for a lipid A modification that leads to the increase of surface charge on the bacterial membrane, acting to repel the positively charged polymyxins as well as positively charged host-derived antimicrobial peptides. We hypothesize that inhibition of ArmR would reverse the resistance to polymyxins, preserving their utility in the clinic, and also sensitize the bacteria to innate host defenses. PUBLIC HEALTH RELEVANCE: Resistance of Gram-negative bacterial pathogens to antibiotics is an increasing and severe healthcare problem. The aim of this research is to better understand the mechanism of Acinetobacter baumannii resistance to last line antibiotics, and to develop new drugs to treat such infections. Since the resistance protein targeted here is widely conserved and present in other drug-resistant bacteria, this work may lead to a novel broad-spectrum antibiotic to combat the most extensively drug-resistant Gram-negative pathogens.